Kaempf Stefanie, Walter Peter, Salz Anna Katharina, Thumann Gabriele
Department of Ophthalmology and IZKF "BIOMAT.", RWTH Aachen University, Pauwelsstr. 30, 52074 Aachen, Germany.
J Neurosci Methods. 2008 Aug 15;173(1):47-58. doi: 10.1016/j.jneumeth.2008.05.018. Epub 2008 Jul 15.
The purpose of this study was to assess survival of adult mammalian neurosensory retina cultured in contact with the layer of a choroid-retinal pigment epithelium (RPE) explant.
The entire adult porcine neurosensory retina and RPE-choroid layer were placed in tissue culture by juxtaposing both tissues in their original orientation. Culture of the neurosensory retina alone and freshly prepared retina were used as control. After 3 days in culture retinal explants were fixed and processed for immunohistochemistry and TUNEL technique.
We observed limited nuclei loss and significant reduction in apoptotic cells in nuclear cell layers (GCL, INL, and ONL) and decreased Muller cell hypertrophy in retina-RPE cultures compared to retinal cultures alone. In addition, cultures were characterized by reduced upregulation of GFAP, vimentin as well as S100 and increased glutamine synthetase expression.
As any tissue culture model, retinal tissue culture is a short-term system and since degenerative processes begin quite early it may be a good model to investigate degenerative processes in the retina. However, our model of culture of retina adjacent to the RPE-choroid layer improves the maintenance of neural retina as evidenced by reduced apoptosis in nuclear cell layers (GCL, INL, and ONL) and reduced gliosis as indicated by the diminished expression of glial-specific proteins and increased glutamine synthetase compared to cultures of retina alone. Thus the retina-RPE-choroid culture system can enable the evaluation of interactions between RPE and neural retina, the role of signaling molecules as well the effect of pharmaceuticals on retinal biology.
本研究的目的是评估与脉络膜视网膜色素上皮（RPE）外植体层接触培养的成年哺乳动物神经感觉视网膜的存活情况。
将整个成年猪神经感觉视网膜和RPE - 脉络膜层以其原始方向并置，放入组织培养中。单独培养神经感觉视网膜和新鲜制备的视网膜作为对照。培养3天后，将视网膜外植体固定并进行免疫组织化学和TUNEL技术处理。
与单独的视网膜培养相比，我们观察到视网膜 - RPE培养物中核细胞层（神经节细胞层、内核层和外核层）的细胞核损失有限，凋亡细胞显著减少，穆勒细胞肥大也减少。此外，培养物的特征是胶质纤维酸性蛋白、波形蛋白以及S100的上调减少，谷氨酰胺合成酶表达增加。
与任何组织培养模型一样，视网膜组织培养是一个短期系统，并且由于退化过程很早就开始了，它可能是研究视网膜退化过程的良好模型。然而，我们的视网膜与RPE - 脉络膜层相邻的培养模型改善了神经视网膜的维持，这表现为核细胞层（神经节细胞层、内核层和外核层）的凋亡减少，以及与单独的视网膜培养相比，胶质特异性蛋白表达减少和谷氨酰胺合成酶增加所表明的神经胶质增生减少。因此，视网膜 - RPE - 脉络膜培养系统能够评估RPE与神经视网膜之间的相互作用、信号分子的作用以及药物对视网膜生物学的影响。
